Bottom Line:
The coiled coil domain of Beclin 1 serves as an interaction platform for assembly of distinct Atg14L- and UVRAG-containing complexes to modulate VPS34 activity.Atg14L and UVRAG promote the transition of metastable homodimeric Beclin 1 to heterodimeric Beclin1-Atg14L/UVRAG assembly.These results suggest that specific utilization of the dimer interface and modulation of the homodimer-heterodimer transition by Beclin 1-interacting partners may underlie the molecular mechanism that controls the formation of various Beclin1-VPS34 subcomplexes to exert their effect on an array of VPS34-related activities, including autophagy.

ABSTRACTBeclin 1 is a core component of the Class III Phosphatidylinositol 3-Kinase VPS34 complex. The coiled coil domain of Beclin 1 serves as an interaction platform for assembly of distinct Atg14L- and UVRAG-containing complexes to modulate VPS34 activity. Here we report the crystal structure of the coiled coil domain that forms an antiparallel dimer and is rendered metastable by a series of 'imperfect' a-d' pairings at its coiled coil interface. Atg14L and UVRAG promote the transition of metastable homodimeric Beclin 1 to heterodimeric Beclin1-Atg14L/UVRAG assembly. Beclin 1 mutants with their 'imperfect' a-d' pairings modified to enhance self-interaction, show distinctively altered interactions with Atg14L or UVRAG. These results suggest that specific utilization of the dimer interface and modulation of the homodimer-heterodimer transition by Beclin 1-interacting partners may underlie the molecular mechanism that controls the formation of various Beclin1-VPS34 subcomplexes to exert their effect on an array of VPS34-related activities, including autophagy.

f1: Structure of the Beclin 1 CC Domain and its imperfect dimer interface.(a) Alignment of the Beclin 1 CC domain sequences from various species. The coloured segments and the italic abcdefg on top of the sequence indicate the heptad repeats typical of CC structure. The coloured spheres below the sequence indicate the residues at a (orange) and d (blue) position of each heptad repeat. (b) The antiparallel dimeric CC structure of the Beclin 1 CC domain. Each monomer helix is coloured in gradient varying from blue at N-terminal to red at C-terminal. The italic letters N and C indicate the N- and C-terminal ends. The dot in the centre marks the two-fold symmetry in the CC domain structure. (c) Helical wheel presentation of the 13 heptad repeats in Beclin 1 CC domain. Only 7 heptad repeats are mapped, representing the unique half of the antiparallel symmetric dimer. (d) Close-up view of the canonical and imperfect a-d' pairings. Each residue is illustrated in ball-and-stick model. Main-chain atoms are coloured blue and side-chain atoms are coloured pink for canonical pairings or golden for imperfect pairings. The a-d' packing is illustrated by van der Waals spheres depicting the side-chain atoms. (e) The distribution of electrostatic charges on Beclin 1 CC domain surface. Colouring scheme is according to GRASP41. The oval circle highlights the E224a-D221e' repulsive pair.

Mentions:
The CC domain of Beclin 1 shares ∼25% sequence similarity with myosin-like proteins, showing a leucine-rich heptad-repeat motif as detonated by 'abcdefg' (Fig. 1a)27. This sequence feature usually induces an oligomeric CC structure, where nonpolar residues at positions a and d form the hydrophobic interface between pairs of helices. The structure of the Beclin 1 CC domain (residue 174–266) was determined by MIRAS using I and Au as two heavy-metal derivatives (Supplementary Table S1 and Supplementary Fig. S2). The Beclin 1 CC domain forms a non-crystallographic dimer as a long antiparallel left-handed CC, with each peptide chain consisting of 13 heptad repeats (Fig. 1b). The first heptad repeat (residues 175–181) of one peptide chain packs against the 13th heptad repeat of the other peptide chain (residues 259–265). Under this arrangement, the dimeric Beclin 1 CC domain structure has an internal two-fold symmetry at the seventh heptad repeat (residue 217-223), where residues 217 and 224 on the two helices are packed against each other. The overall geometry of the Beclin 1 CC domain structure conforms to that of the classic CC (Supplementary Fig S3). The full length of the CC domain is ∼129 Å, which roughly comprises one helical pitch made up of the two symmetry-related halves (Fig. 1b).

f1: Structure of the Beclin 1 CC Domain and its imperfect dimer interface.(a) Alignment of the Beclin 1 CC domain sequences from various species. The coloured segments and the italic abcdefg on top of the sequence indicate the heptad repeats typical of CC structure. The coloured spheres below the sequence indicate the residues at a (orange) and d (blue) position of each heptad repeat. (b) The antiparallel dimeric CC structure of the Beclin 1 CC domain. Each monomer helix is coloured in gradient varying from blue at N-terminal to red at C-terminal. The italic letters N and C indicate the N- and C-terminal ends. The dot in the centre marks the two-fold symmetry in the CC domain structure. (c) Helical wheel presentation of the 13 heptad repeats in Beclin 1 CC domain. Only 7 heptad repeats are mapped, representing the unique half of the antiparallel symmetric dimer. (d) Close-up view of the canonical and imperfect a-d' pairings. Each residue is illustrated in ball-and-stick model. Main-chain atoms are coloured blue and side-chain atoms are coloured pink for canonical pairings or golden for imperfect pairings. The a-d' packing is illustrated by van der Waals spheres depicting the side-chain atoms. (e) The distribution of electrostatic charges on Beclin 1 CC domain surface. Colouring scheme is according to GRASP41. The oval circle highlights the E224a-D221e' repulsive pair.

Mentions:
The CC domain of Beclin 1 shares ∼25% sequence similarity with myosin-like proteins, showing a leucine-rich heptad-repeat motif as detonated by 'abcdefg' (Fig. 1a)27. This sequence feature usually induces an oligomeric CC structure, where nonpolar residues at positions a and d form the hydrophobic interface between pairs of helices. The structure of the Beclin 1 CC domain (residue 174–266) was determined by MIRAS using I and Au as two heavy-metal derivatives (Supplementary Table S1 and Supplementary Fig. S2). The Beclin 1 CC domain forms a non-crystallographic dimer as a long antiparallel left-handed CC, with each peptide chain consisting of 13 heptad repeats (Fig. 1b). The first heptad repeat (residues 175–181) of one peptide chain packs against the 13th heptad repeat of the other peptide chain (residues 259–265). Under this arrangement, the dimeric Beclin 1 CC domain structure has an internal two-fold symmetry at the seventh heptad repeat (residue 217-223), where residues 217 and 224 on the two helices are packed against each other. The overall geometry of the Beclin 1 CC domain structure conforms to that of the classic CC (Supplementary Fig S3). The full length of the CC domain is ∼129 Å, which roughly comprises one helical pitch made up of the two symmetry-related halves (Fig. 1b).

Bottom Line:
The coiled coil domain of Beclin 1 serves as an interaction platform for assembly of distinct Atg14L- and UVRAG-containing complexes to modulate VPS34 activity.Atg14L and UVRAG promote the transition of metastable homodimeric Beclin 1 to heterodimeric Beclin1-Atg14L/UVRAG assembly.These results suggest that specific utilization of the dimer interface and modulation of the homodimer-heterodimer transition by Beclin 1-interacting partners may underlie the molecular mechanism that controls the formation of various Beclin1-VPS34 subcomplexes to exert their effect on an array of VPS34-related activities, including autophagy.

ABSTRACTBeclin 1 is a core component of the Class III Phosphatidylinositol 3-Kinase VPS34 complex. The coiled coil domain of Beclin 1 serves as an interaction platform for assembly of distinct Atg14L- and UVRAG-containing complexes to modulate VPS34 activity. Here we report the crystal structure of the coiled coil domain that forms an antiparallel dimer and is rendered metastable by a series of 'imperfect' a-d' pairings at its coiled coil interface. Atg14L and UVRAG promote the transition of metastable homodimeric Beclin 1 to heterodimeric Beclin1-Atg14L/UVRAG assembly. Beclin 1 mutants with their 'imperfect' a-d' pairings modified to enhance self-interaction, show distinctively altered interactions with Atg14L or UVRAG. These results suggest that specific utilization of the dimer interface and modulation of the homodimer-heterodimer transition by Beclin 1-interacting partners may underlie the molecular mechanism that controls the formation of various Beclin1-VPS34 subcomplexes to exert their effect on an array of VPS34-related activities, including autophagy.